Wataru Makino

Extreme population genetic differentiation and secondary contact in the freshwater copepod Acanthodiaptomus pacificus in the Japanese Archipelago

We investigated the sequence variation in the mitochondrial cytochrome c oxidase subunit 1 (mtCOI) gene and the nuclear ribosomal internal transcribed spacers (ncITS) of the calanoid copepod Acanthodiaptomus pacificus in Japan. A. pacificus individuals were divided into three divergent mtCOI lineages (mt‐A, ‐B and ‐C). mt‐A was distributed in the northernmost part of Japan, from Hokkaido to the northern part of Honshu Island, whereas mt‐C was the southernmost lineage, distributed from central Honshu to Shikoku and Kyushu Islands. mt‐B was distributed between these former two lineages, resulting in parapatry with mt‐C and mt‐A. In all lineages, 80% of the localities were fixed for a single haplotype, and different localities tended to have different haplotypes. The degree of genetic differentiation among these lineages (15–22%) was at an interspecific level, according to the criteria of the DNA barcode technique. However, the topology of ncITS was not congruent with that of mtCOI, as the reciprocal monophyly was not observed within mt‐B and mt‐C. Therefore, we merged them into the Southern Lineage and separated it from the Northern Lineage (i.e. mt‐A). Evidence of introgression was found within the Southern Lineage, while gene flow was not observed between the Northern and Southern Lineages, suggesting that A. pacificus is a cryptic species complex. We also argue that genetic differentiations of A. pacificus in Japan may reflect the history of separation, transgression and regression of the landmass during the formation of current Japanese Archipelago.

The long-term consequences of hybridization between the two Daphnia species, D. galeata and D. dentifera, in mature habitats

BackgroundEcological specializations such as antipredator defense can reinforce morphological and distributional divergence within hybridizing species. Two hybridizing species of Daphnia (D. galeata and D. dentifera) are distributed in both Japan and North America; however, these populations have a longer history in Japan than in North America due to the differing impact of the last glaciation on these two regions. We tested the hypothesis that this longer coexistence in Japan would lead to extensive genetic admixture in nuclear and mitochondrial DNA whilst the distinct morphological traits and distributional patterns would be maintained.ResultsThe high level of correspondence among morphological traits, distribution, and mitochondrial and nuclear DNA types for the specimens with D. dentifera mtDNA indicated that the species distinction has been maintained. However, a discordance between mtDNA and nuclear ITS-1 types was observed for most specimens that had D. galeata mtDNA, consistent with the pattern seen between the two species in North America. This observation suggests nuclear introgression from D. dentifera into D. galeata without mitochondrial introgression.ConclusionsThe separation of morphological traits and distribution ranges of the two hybridizing species in Japan, as well as in North America, has been maintained, despite large differences in climatic and geographical histories of these two regions. Variations in environmental factors, such as predation pressure, might affect maintenance of the distribution, although the further studies are needed to confirm this.

Shifts in morphology and diet of non-native sticklebacks introduced into Japanese crater lakes.

An increasing number of exotic animals are causing ecological problems. Therefore, for better ecosystem management, it is important to understand how exotic species colonize and adapt to novel environments. The threespine sticklebacks (Gasterosteus aculeatus) can be a good vertebrate model system to explore the ecological and genetic mechanisms of adaptation not only in natural populations, but also in non-native populations. Although morphological changes have been documented in several introduced populations of stickleback, little is known about the dietary changes during colonization into novel environments. Here, we investigated the morphological and dietary changes of exotic threespine stickleback populations introduced into three Japanese crater lakes (Lake Towada, Lake Kussharo, and Lake Shikotsu). Sticklebacks were introduced into the crater lakes likely along with salmonids transplanted for aquaculture. The stickleback population in Lake Kussharo had multiple mitochondrial haplotypes and had larger phenotypic variances than other crater lake stickleback populations that had only one mitochondrial haplotype. Compilation of historical data on the morphology and stomach contents of the Lake Towada stickleback population showed that substantial shifts in body size and stomach contents occurred after colonization. Some of these changes may be related to an outbreak of the Schistocephalus parasite. These results suggest that sticklebacks can change their morphology and trophic ecology when they colonize novel environments. Therefore, extreme care should be taken when salmonids are transported between watersheds for aquaculture and that long-term monitoring of exotic species is essential for ecosystem management. In addition, further genetic studies on phenotypic changes in crater lake sticklebacks would help elucidate the genetic mechanisms underlying the adaptation of exotic fishes to novel environments.

DNA extraction and amplification methods for ephippial cases of Daphnia resting eggs in lake sediments: a novel approach for reconstructing zooplankton population structure from the past

This study describes a method of DNA extraction and amplification for ephippial cases of Daphnia resting eggs from lake sediment. Recent studies have reconstructed succession records of Daphnia species by genetically analyzing Daphnia resting eggs stored in lake sediments and revealed changes in dominant Daphnia species that correspond well with environmental changes. However, this approach is not applicable to lakes where most of the resting eggs in the sediment have already hatched out. We modified conventional methods for DNA extraction and amplification to enable genetic analyses of the ephippial case that envelops and protects the resting eggs, and we compared the performance of the modified method to the conventional one. We confirmed that we could efficiently analyze the sequences of the ephippial cases collected in a sediment core using our modified method. It enables us to reconstruct changes in genetic structure of Daphnia populations regardless of hatching rates of the resting eggs.

Stoichiometric effects of warming on herbivore growth: experimental test with plankters

Recent studies have shown that putative warming increases water temperatures in many lakes. A higher temperature would directly affect the growth rates of lake organisms; however, the growth responses of lake herbivores to increased temperatures may be complex. Under limited nutrient supplies, for example, increased temperature may accelerate the growth of phytoplankton but consequently decrease their nutrient content, which may in turn reduce herbivore growth. Thus, temperature-driven declines in food nutrient content may offset the positive effects of warming on herbivore growth depending on the trophic condition of habitats. We examined this hypothesis by laboratory experiments that simultaneously manipulated temperature and stoichiometric interactions between green algae (Scenedesmus) and a planktonic herbivore (Daphnia) under various phosphorous conditions. The following experiments were performed with Scenedesmus and Daphnia at the same temperatures (12, 16, 20 and 24°C). In the first experiment, Scenedesmus were grown in semi-batch cultures at four different phosphorus supply rates with a fixed dilution rate (i.e., algal loss rate). In the second experiment, algae were grown at four different dilution rates with a fixed phosphorus supply level. Then, the steady-state algae were harvested daily and fed directly to juvenile Daphnia to measure their specific growth rates in both experiments. Increased temperature accelerated Daphnia growth in both experiments. However, the effect of temperature decreased with decreasing phosphorus supply rate and dilution rates, although algae were sufficiently abundant. This was because the algal phosphorus:carbon ratio decreased with increasing temperature, especially when phosphorus availability for algae was small due to low phosphorus supply or dilution rates. The results clearly showed for the first time that direct stimulation of herbivore growth by warming can be offset by poor food stoichiometry. We suggest that in lakes with low algal growth rates due to low nutrient inputs or in lakes with low algal loss rates due to a shortage of algal grazers, warming may indirectly slow herbivore growth despite increased algal biomass.

Cryptic diversity of Japanese Diaphanosoma (Crustacea: Cladocera) revealed by morphological and molecular assessments

Abstract We found 5 distinct species of Diaphanosoma from a variety of lakes in Japan according to morphological examination and genetic analyses with the DNA sequences of mitochondrial 16S ribosomal RNA and cytochrome c oxidase subunit I genes (mtCOI). Previously, D. brachyurum was thought to occur at temperate regions including Japan; however, we did not find D. brachyurum in Japan. Instead, we found D. cf. amurensis, which inhabited mainly natural habitats of high altitude and latitude, and D. cf. dubium, D. cf. orientalis, and D. cf. macrophthalma, which occurred in a variety of habitats from large natural lakes to small artificial ponds at low altitude. We also found another species from only one pond that possessed mtCOI sequences that matched almost completely with those from North American specimens and thus was probably nonindigenous Diaphanosoma. Concordance between the results of morphological identifications and genetic analyses showed that the DNA barcodes established in this study are useful to identify Diaphanosoma species in Japan and adjacent areas.

Ecological and genetic impact of the 2011 Tohoku Earthquake Tsunami on intertidal mud snails

Natural disturbances often destroy local populations and can considerably affect the genetic properties of these populations. The 2011 Tohoku Earthquake Tsunami greatly damaged local populations of various coastal organisms, including the mud snail Batillaria attramentaria, which was an abundant macroinvertebrate on the tidal flats in the Tohoku region. To evaluate the impact of the tsunami on the ecology and population genetic properties of these snails, we monitored the density, shell size, and microsatellite DNA variation of B. attramentaria for more than ten years (2005–2015) throughout the disturbance event. We found that the density of snails declined immediately after the tsunami. Bayesian inference of the genetically effective population size (Ne) demonstrated that the Ne declined by 60–99% at the study sites exposed to the tsunami. However, we found that their genetic diversity was not significantly reduced after the tsunami. The maintenance of genetic diversity is essential for long-term survival of local populations, and thus, the observed genetic robustness could play a key role in the persistence of snail populations in this region which has been devastated by similar tsunamis every 500–800 years. Our findings have significant implications for understanding the sustainability of populations damaged by natural disturbances.

Historical changes in the ecosystem condition of a small mountain lake over the past 60 years as revealed by plankton remains and Daphnia ephippial carapaces stored in lake sediments.

To examine if changes in species composition of a plankton community in the past due to anthropogenic activities can be clarified in lakes without any monitoring data, we analyzed genetically ephippial carapaces of Daphnia with plankton remains stored in the bottom sediments of Lake Hataya Ohunma in Japan. In the lake, abundance of most plankton remains in the sediments was limited and TP flux was at low levels (2–4 mg/m2/y) before 1970. However TP flux increased two-fold during the period from 1980s to 1990s. In parallel with this increase, abundance of most plankton remains increased although abundance of benthic testate amoebae’s remains decreased, indicating that the lake trophic condition had changed from oligo- to mesotrophic for the past 60 years. According to cluster analysis, the stratigraphic sediments were divided into two periods with different features of the phytoplankton composition. Chronological comparison with events in the watershed suggested that eutrophication occurred because of an increase in visitors to the watershed and deposition of atmospheric dust. In this lake more than 50% of resting eggs produced by Daphnia over the past 60 years hatched. However, genetic analysis of the ephippial carapaces (remains) showed that the Daphnia population was originally composed of D. dentifera but that D. galeata, or its hybrid with D. dentifera, invaded and increased the population density when the lake was eutrophied. Subsequently, large D. pulex established populations in the 1980s when largemouth bass were anonymously introduced. These results indicated that the Lake Hataya Ohunma plankton community underwent significant changes despite the fact that there were no notable changes in land cover or land use in the watershed. Since increases in atmospheric deposition and release of fish have occurred in many Japanese lakes, the changes in the plankton community described here may be widespread in these lakes.

Characterization and cross-species utility of polymorphic microsatellite markers in the Asian mud snails Batillaria attramentaria and B. multiformis

We describe 16 polymorphic microsatellite markers in two congenic species of mud snails, Batillaria attramentaria and B. multiformis, as well as the conditions of multiplexing and the cross-species availability of these loci. The number of alleles per locus ranged from 2 to 8 for B. attramentaria and from 2 to 21 for B. multiformis. The set of markers described here will constitute efficient tools for detecting the loss of their genetic diversity due to the tsunami disturbance of the 2011 Tohoku Earthquake.

Food quality determinants for Daphnia growth in P-limited lakes

The growth ofherbivorous zooplankton is crucial for production at higher trophic levels in lakes. Among the environmental factors affecting growth rates of planktonic herbivores, food availabiliry has received much attention because the abundance and production of phytoplankton vary gready in nature. In addition, growing evidence suggests that zooplankton grazers potentially suffer nor only from insufficient food quantity but frequently also from inadequate food quality (ELSER er al. 2000, HESSEN & FAAFENG 2000). While a large number of studies have addressed food quantity effects on herbivore growth, the relative importance of food quality and quantity has rarely been assessed in nature (STERNER & SCHULZ 1998). Other than toxiciry, size and morphology, relarive contents ofN, P and some bio-chemicals have been proposed as determinants of food quality for herbivores (HESSEN 1992, URABE & WATANABE 1992, MüLLER-NAvARRA 1995a, AHLGREN et al. 1997, .K!LHAM et al. 1997). Much of this work has focused on Daphnia, a key herbivore in lake food webs (CARPENTER & .K!TCHELL 1993). MDLLER-NAVARRA (1995a) showed that Daphnia growth in a German lake was most strongly correlated with the content of fatty acids, especially eicosapentaenoic acid (20:5ro3, EPA) in seston. In contrast, ELSER et al. (2001) demonstrated that P amendment to seston from P-limited lakes stimulated Daphnia growth, independent of any changes in fatty acids. These inconsistent results suggest that the nutritional substance determining food quality for Daphnia growth is site specific. If this is the case, it is important to determine when and where a particular nutritional substance is involved in determining food quality (STERNER 1997). In the present study, somatic growth rates of Daphnia individuals were measured using seston from P-limited lakes and from manipulated enclosures experiencing different light and P-supply regimes. Since production and chemical composition of algae vary widely depending on light and nutrient regimes (URABE & STERNER 1996, STERNER et al. 1998), field enclosures with different light and nutrient regimes provided a good opportunity to examine the effects of various nutritional substances in food on Daphnia growth. Specific goals were to evaluate (I) the relative importance of food quality and quantity, and (2) the relative importance of various putative substances in determining food quality for Daphnia growth.